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Systems for Dramatically Improved Synthetic RNA

显着改进合成 RNA 的系统

基本信息

批准号：
10557074
负责人：
Craig T Martin
金额：
$ 30.63万
依托单位：
UNIVERSITY OF MASSACHUSETTS AMHERST
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-04-01 至 2025-01-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10557074
关键词：
Adoption Affinity Affinity Chromatography Area Basic Science Binding Biochemistry Biological Assay Biology Biophysics Biotechnology Cellular biology Complementary DNA Creativeness DNA Dependence Development Discipline Double-Stranded RNA Enzymes Feedback Fostering Gel Genetic Transcription Guide RNA Health Immobilization In Vitro Innate Immune Response Intervention Length Life Messenger RNA Methods Mission Modeling Molecular Biology Nanotechnology Nature Pathway interactions Performance Polymerase Primer Extension Public Health RNA RNA Sequences RNA chemical synthesis Randomized Reaction Research Research Personnel Resistance Running Sodium Chloride Surveys System T7 RNA polymerase Technology Testing Therapeutic Time United States National Institutes of Health Work catalyst design high standard improved innovation novel strategies prevent promoter success synthetic biology therapy design tool transcriptome sequencing

项目摘要

Project Summary A wide variety of RNA researchers synthesize their RNA using the enzyme T7 RNA polymerase, as this enzyme is robust and can yield large quantities of RNA, at any length scale. However, undesired products typically contaminate the desired RNA, in often unpredictable ways. This is perhaps most impactful currently in the mRNA therapeutics field, where contaminating double stranded RNAs can trigger a potentially lethal innate immune response, but contaminants almost certainly impact other studies as well, from basic research in cell and molecular biology and biochemistry/biophysics, to synthetic biology, to RNA nanotechnology. Gel purifications are tedious, low yield, and imperfect (indeed, the darkest band on the gel may not be the correct product, and even the correct length RNA pool can be heterogeneous!). Longer RNA impurities derive from correct RNA, reducing yields at synthesis, and can be distributed across a wide range of lengths, making gel analysis problematic. Building on new mechanistic understandings, this project will develop systems that limit the conditions that give rise to these impurities, specifically by inhibiting the off-pathway reactions, and will develop simple affinity purification approaches – all with an aim towards achieving monodisperse RNAs of defined length and sequence. Sensitive analytical and functional assays for success will be developed and applied to guide design. Tools will be developed with an eye towards broad adoption by a variety of researchers.

项目总结